Abstract 315 Poster Session IV, Tuesday, 5/4 (poster 323)
We have recently isolated a receptor for SP-A (SP-AR) specifically expressed by type II cells (TIIC). The goals of this study were to determine the ontogeny of SP-AR protein levels, and if endogenous substances within the lung, mammalian bombesin (B) and tRA, regulate SP-AR protein levels. Methods: Whole lungs from 18d, 20d, 22d fetal, newborn, 1-14d postnatal, and adult rats were homogenized, and levels of SP-AR were analyzed by immunoblot using polyclonal antibody to SP-AR. An ELISA with monoclonal antibody to SP-AR was performed on primary cultures of TIIC from fetal and adult rats. The ligand, SP-A, was used to block the binding of antibody to determine specific binding of antibody to SP-AR. Explants of 16d fetal rat lung were incubated in media (control) or media with 1 µM B for 0-48h. Explants of 21d fetal rat lung were incubated in media+/- 1-100 nM tRA for 4h to 7d. Tissue was harvested and SP-AR protein levels were analyzed by immunoblot. Results: There was an increase in detectable SP-AR in whole lung between 18d and 22d in fetal rat, a further increase from 22d to 1d postnatal age, a decrease from 2d to 7d, followed by an increase at 14d and adult rat lung. There was minimal binding of SP-AR antibody to fetal TIIC with maximal binding (Bmax) that was 10% of adult values. Binding was saturable and specific in adult TIIC. The ligand, SP-A, completely blocked binding of antibody in adult cells but not in fetal cells (table).
B increased levels of SP-AR after 6-48h exposure in 16d fetal rat lung explants. This effect was associated with increased cellular proliferation as measured by changes in Proliferating Cell Nuclear Antigen in both explants and fetal TIIC. SP-AR levels in 21d fetal rat lung explants increased after 4h and 1d exposures to 1 nM and 100 nM tRA, respectively. There was a progressive decline in this effect with time of incubation, so that by 5-7d there was inhibition of SP-AR levels by tRA. Conclusions: We conclude that SP-AR number increases during the saccular phase (late fetal life), declines in the early alveolar phase (2-7d postnatal) and then increases again in the late alveolar phase of lung development (14d after birth to adult). Receptor Bmax is minimal in fetal TIIC and thus no blockade by the ligand is detectable. Binding is specific and saturable in adult TIIC. B enhances SP-AR levels in fetal rat lung and stimulates TIIC proliferation. There is a biphasic effect of tRA on SP-AR similar to the effect of tRA on SP-A, SP-B and SP-C that has been reported.